Inulin Might Exceed Incretin Based Drugs in the Treatment of type 2 Diabetes Mellitus

1Lecturer of Internal Medicine, Endocrinology, Diabetology & Metabolism Medical Department, Medical Division, National Research Centre (NRC), Cairo, Egypt. 2Professor of Internal Medicine, Endocrinology and Metabolism, Department of Internal Medicine, Medical Division, National Research Centre (NRC), Cairo, Egypt. 3Professor of Nutrition-Department of Nutrition and Food Science, National Research Centre (NRC), Cairo, Egypt. 4Professor of Biochemistry-Medical Biochemistry Department, Medical Division, National Research Centre (NRC), Cairo, Egypt. 5Professor of Biochemistry-Medical Biochemistry Department, Medical Division, National Research Centre (NRC), Cairo-Egypt. 6Assisstant Professor of Biochemistry-Medical Biochemistry Department, Medical division, National Research Centre (NRC), Cairo-Egypt. *Corresponding author E-mail: mwgnicola@hotmail.com

The pancreatic endocrine portion (islets of Langerhans) is strongly involved in the regulation of energy metabolism. These islets comprise four kinds of hormone secreting cells; the á cells secrete glucagon, the â cells secrete insulin and amylin. The ä cells secrete somatostatin while the fourth type of cells secrete the pancreatic polypeptide 1 .
Insulin and glucagon are the two main hormones which are largely responsible for blood glucose and energy homeostasis 2 .
Insulin is a hypoglycemic and anabolic hormone. It acts on the liver, muscles and adipose tissue. It enhances glucose uptake by insulin sensitive cells and its conversion into glycogen (in liver & muscles) and to fat (in adipose tissue), besides it has an anabolic effect on protein 3 .
Glucagon, on the other hand acts in an opposite manner to insulin. It is catabolic and hyperglycemic. It activates glycogenolysis and gluconeogenesis mainly in the liver, also it inhibits glycogenesis and glycolysis in addition to its lipolytic action on the adipose tissue 3 . The glycemic effect of glucagon is essential to regain and maintain the blood glucose level in hypoglycemic states. This is very important to vital organs as the brain which is dependant on blood glucose as its main fuel 4 .
In normal glycemic states, both insulin and glucagon hormones cooperate in maintaining the blood glucose level in the normal range 2,5 . In diabetes mellitus, there is a disturbance in glucagon/insulin secretion. There is an abnormal increase in blood glucagon level relative to insulin level 3 .
Inulin a naturally occuring oligosaccharide of plant origin, has been known to have hepatic lipotropic and blood glucose lowering effect 6,7 . It passes undigested in the small intestine to the colon, where it is acted upon by certain strains of the colonic microbiome. These secrete peptides which stimulate the L-endocrine cells of the small bowel to produce glucagon like peptide-1 (GLP-1) 8,9 . This incretin hormone suppresses glucagon and stimulates insulin secretion 10,11 .
In this study, we aim to find out, how inulin ingestion affects blood glucagon level and how inulin can participate in the regulation of blood glucose level and glycemic contol.

Subjects
Twenty eight obese, type 2 diabetic women were recruited from the municipal hospitals of Cairo, Egypt. The age range was 40-65 years.
Inclusion criteria: obese (body mass index > 30), type 2 diabetic women, middle aged and above, hypertensive or not and not under antilipidemic drugs.

Exclusion criteria
Hormonal therapy or contraceptive pills. Endocrine disorders or other metabolic diseases.
Malignancy or organ failure (lung, heart, liver or Kidney). Local or systemic infection including chest, urinary tract, gastro-intestinal tract or skin infection (local or extensive).

Ethical committee approval
The present work has been approved by the National Research Centre (NRC) committee of ethics, Cairo, Egypt. (Certificate number 15011).

Consent
All patients signed consent for participation.

METhodS
Prebiotic inulin was taken by each of the individuals of the studied group as an add on therapy to their conventional ongoing antidiabetic treatment. Four grams of inulin were given with milk to each patient daily; (half of the dose in the morning and the other half in the evening) for three weeks. This dose was chosen empirically putting in mind that a small amount of inulin can exert a bifidogenic effect 12 .
Each patient was subjected before and after the period of inulin intake for the following. A-Full history. B-Thorough clinical examination. C-Laboratory investigations:

Laboratory investigations Estimation of fasting serum glucagon
Serum glucagon was estimated by ELISA technique after Bak et al. 13 . Human GC (Glucagon) ELISA Kit is used from Elabscience. Catalogue number: E-EL-H2237 96T. Elabscience Biotechnology Co., Ltd. Website: www.elabscience. com.

determination of insulin resistance (hoMA-IR)
Insulin resistance (IR) was assessed using the equation of homeostasis model assessment for insulin resistance (HOMA-IR) utilizing fasting serum glucose & fasting serum insulin values, after Mathiews et al. 14 .

Fasting serum insulin determination
This was quantitatively determined using an enzyme immunoassay method according to the National Committee for Clinical Laboratory Standards 15 . Manufacturer of kit used: immunospect

Fasting serum glucose assessment
This was assessed by an enzymatic colorimetric method. This is an enzymatic oxidation method for glucose by glucose oxidase enzyme, according to Tietz 16 .

Statistical methods
Results were coded, tabulated and statistically analyzed. These were done using IBM Statistical Package for Social Sciences (SPSS statistics) software version 22.0; IBM Corp. Chicago-USA-2013.
Quantitative data were described statistically as minimum and maximum for the range as well as the mean± standard deviation (SD) for quantitative parametric values. The paired t-test was used for inferential analysis of the quantitative variables in cases of two dependent groups having parametric data. Significance level was taken as P value < 0.05.

RESULTS
There was significant decrease in mean fasting serum level of glucagon after inulin ingestion period from 188.3±50.3 to 164.9±46.9 with P value <0.001, table [1] and figure [1].
There was significant decrease in mean fasting serum value of HOMA-IR after inulin intake period from 11.1±8.2 to 7.2±5.0 with P value <0.001, table [1] and figure [2].

dISCUSSIoN
The glucagon/insulin ratio is a fundamental factor in the control and regulation of hepatic glucose metabolism. In normoglycemic subjects, glucagon secretion by pancreatic á cells is inhibited by blood glucose levels lower than those needed for â cell stimulation and insulin secretion 17,18 . In diabetics, on the other hand, there is relative increase in the glucagon/insulin ratio 3 . However the disturbed glucagon/insulin ratio is not the only disorder met with. Actual hyperglucagonemia may occur. This increase in glucagon secretion is one of the main factors behind the insulin resistance and the hyperglycemic state in diabetes mellitus 19,20 .
The present work has shown a positive correlation between the fasting serum glucagon level and the insulin resistance as estimated by HOMA-IR equation in the studied diabetic patients. Also, in the present study, inulin ingestion significantly decreased both fasting serum glucagon level and insulin resistance manifested by lowered HOMA-IR.
In fact, inulin has got no direct action on both á and â cells of the islets of Langerhans in the pancreas. As we mentioned before, it stimulates certain colonic microbacteria. These release particular peptides which when absorbed, reach the endocrine L cells in the small intestine. These L cells release their glucagon like peptide-1 (GLP-1). This incretin suppresses glucagon secretion and enhances insulin release in a glucose dependant manner 10,11 . In this respect inulin acts in a similar way like the novel smart incretin based  21 . These include the GLP-1 analogues or mimetics and the dipeptidyl peptidase type 4 (DPP4) inhibitors. Being glucose dependant in their action, they protect the diabetic patient against the possible hazardous hypoglycemic side effect of antidiabetic therapy such as insulin or insulin secretagogues.
Moreover, in previous studies by the present authors, inulin reduced insulin resistance by lowering serum triglycerides and improving hepatosteatosis. Fatty liver is the prime and main site for insulin resistance 6,7 . Besides, in other previous studies by the present authors, inulin proved to tackle various important facets in the pathogenesis of type 2 diabetes mellitus and its effect on the vascular tree. These studies proved that inulin possesses anti-inflammatory properties 22 ; Also inulin has a protective effect on the microvasculature 23 . In addition to its antiatherogenic effect on the macrovasculature 24 . Taking into consideration the results of this work together with our preceeding studies, 6,7,22-24 inulin might exceed the novel incretin based drugs in the treatment of diabetes mellitus.

CoNCLUSIoN
In conclusion, in the present work, inulin proved to reduce insulin resistance through lowering glucagon hormone level which has an insulin antagonizing action. This together with its triglycerides lowering effect and hepatic lipotropic effect makes inulin a reasonable antinsulin resistant agent. Moreover, as it works in a glucose dependant manner, it can be added safely to the ongoing treatment of diabetic patients. Also, it can be given alone to prediabetics (patients with impaired fasting glucose or those with impaired glucose tolerance).
Again, we would like to draw the attention for critical cases which deserve meticulous study. Sometimes diabetic patients on insulin therapy whether type 1 or long standing type 2, suffer attacks of hypoglycemia due to failure of responsiveness of their á cells to secrete glucagon 25 . This abnormal phenomenon has been attributed to multiple failures in á cell regulations including insulin resistance 26 . The exact underlying causes are still vague. The authors suggest to reduce insulin dose and add inulin as an adjunct therapy to those patients to study such cases.

ACKNoLWdGEMENTS
Thanks for National Research Center, Cairo, Egypt for financing this work. Again, thanks to all patients who volunteered to participate in this study.